• Title/Summary/Keyword: Single-molecule

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Role of N-terminal Hydrophilic Amino Acids in Molecular Translocation of CTLA-4 to Cell Surface (CTLA-4 항원의 세포막 도달 기작에서 친수성 N말단 아미노산 잔기의 역할)

  • Han, Ji-Woong;Lee, Hye-Ja;Kim, Jin-Mi;Choi, Eun-Young;Chung, Hyun-Joo;Lim, Soo-Bin;Choi, Jang-Won;Chung, Yong-Hoon
    • IMMUNE NETWORK
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    • v.2 no.2
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    • pp.102-108
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    • 2002
  • Background: This study was aimed to differentiate two forms of CTLA-4 (CD152) in activated peripheral blood lymphocyte and clarify the mechanism how cytoplasmic form of this molecule is targeted to cell surface. Methods: For this purpose we generated 2 different anti-human CD152 peptide antibodies and 5 different N'-terminal deletion mutant CTLA4Ig fusion proteins and carried out a series of Western blot and ELISA analyses. Antipeptide antibodies made in this study were anti-CTLA4pB and anti-CTLA4pN. The former recognized a region on extracellular single V-like domain and the latter recognized N'-terminal sequence of leader domain of human CD152. Results: In Western blot, the former antibody recognized recombinant human CTLA4Ig fusion protein as an antigen. And this recognition was completely blocked by preincubating antipeptide antibody with the peptide used for the antibody generation at the peptide concentration of 200 ug/ml. These antibodies were recognized human CD152 as a cytoplasmic sequestered- and a membrane bound- forms in phytohemagglutinin (PHA)-stimulated peripheral blood lymphocyte (PBL). These two forms of CD152 were further differentiated by using anti-CTLA4pN and anti-CTLA4pB antibodies such that former recognized cytosolic form only while latter recognized both cytoplasmic- and membraneforms of this molecule. Furthermore, in a transfection expression study of 5 different N'-terminal deletion mutant CTLA4Ig, mutated proteins were secreted out from transfected cell surface only when more than 6 amino acids from N'-terminal were deleted. Conclusion: Our results implies that cytosolic form of CTLA-4 has leader sequence while membrane form of this molecule does not. And also suggested is that at least N'-terminal 6 amino acid residues of human CTLA-4 are required for regulation of targeting this molecule from cytosolic- to membrane- area of activated human peripheral blood T lymphocyte.

Delphinidin Chloride Effects on the Expression of TNF-$\alpha$ Induced Cell Adhesion Molecules (TNF-$\alpha$에 의해 유도된 세포부착분자의 발현에 대한 Delphinidin chloride의 억제 효과)

  • Koh, Eun-Gyeong;Chae, Soo-Chul;Seo, Eun-Sun;Na, Myung-Suk;Lee, Jong-Bin
    • Korean Journal of Environmental Biology
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    • v.27 no.1
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    • pp.88-94
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    • 2009
  • The process of atherosclerosis begins through secretion of inflammatory cytokine or adhesion of leukocyte from damage in blood vessels and transmigration. This study was conducted to investigate the effects of delphinidin chloride (DC) in the initial process of atherosclerosis on the expression of ICAM-1 (Intracellular Adhesion Molecule-1) and VCAM-1 (Vascular Adhesion Molecule-1) related to adhesion of leukocyte at the HUVEC (human umbilical vein endothelial cell line. As a result, cell growth inhibition rate at 50 ${\mu}M$ was respectively 4, 3 and 5% without cell toxicity. As a result of morphological observation monocyte-endothelial cell adhesion assay and optical microscope carried out to measure attachment of mononuclear cells to endothelial cells induced by Tumor necrosis factor-alpha (TNF-$\alpha$) at concentrations without cell toxicity, DC concentration-dependently suppressed attachment. When effects on the expression of VCAM-1 and ICAM-1, cell adhesion molecules induced from endothelial cells by TNF-$\alpha$, were comparatively analyzed using western blot analysis and RT-PCR methods, protein of VCAM-1 and ICAM-1 and expression at the level of mRNA were concentration-dependently reduced. Taken together, the results of this studies provide evidence that DC possess an anti-metastatic activity.

Synthesis and Crystal Structure of Lead Iodide in the Sodalite Cavities of Zeolite A (LTA)

  • Kim, Seok-Han;Lim, Woo-Taik;Kim, Ghyung-Hwa;Lee, Heung-Soo;Heo, Nam-Ho
    • Bulletin of the Korean Chemical Society
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    • v.27 no.5
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    • pp.679-686
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    • 2006
  • The positions of $PbI _2$ molecule synthesized into the molecular-dimensioned cavities of $\mid K_6 (Pb _4I_2)(PbI_2) _{0.67}-(H_2O)_2\mid [Si _{12}Al _{12}O _{48}]$-LTA have been determined. A single crystal of $\mid Pb _6\mid [Si _{12}Al _{12}O _{48}]$-LTA, prepared by the dynamic ion-exchange of $\mid Na _{12}\mid [Si _{12}Al _{12}O _{48}]$-LTA with aqueous 0.05 M $Pb _(NO _3)_2$ and washed with deionized water, was placed in a stream of flowing aqueous 0.05 M KI at 294 K for three days. The resulting crystal structure of the product $( \mid K_6 (Pb _4I_2)(PbI_2) _{0.67}(H_2O)_2\mid [Si _{12}Al _{12}O _{48}]$-LTA, a = 12.353(1) $\AA$) was determined at 294 K by single-crystal X-ray diffraction in the space group Pm3 m. It was refined with all measured reflections to the final error index $R_1$ = 0.062 for 623 reflections which $F_o$ > 4$\sigma$($F_o$). 4.67 $Pb ^{2+}$ and six $K^+$ ions per unit cell are found at three crystallographically distinct positions: 3.67 $Pb ^{2+}$ and three $K^+$ ions on the 3-fold axes opposite six-rings in the large cavity, three $K^+$ ions off the plane of the eight-rings, and the remaining one $Pb ^{2+}$ ion lies opposite four-ring in the large cavity. 0.67 $Pb ^{2+}$ ions and 1.34 $I^-$ ions per unit cell are found in the sodalite units, indicating the formation of a $PbI _2$ molecule in 67% of the sodalite units. Each $PbI _2$ (Pb-I = 3.392(7) $\AA$) is held in place by the coordination of its one $Pb ^{2+}$ ion to the zeolite framework (a $Pb ^{2+}$ cation is 0.74 $\AA$ from a six-ring oxygens) and by the coordination of its two $I^-$ ions to $K^+$ ions through six-rings (I-K = 3.63(4) $\AA$). Two additional $I^-$ ions per unit cell are found opposite a four-ring in the large cavity and form $Pb _2K_2I^{5+}$ and $Pb _2K_2I^{3+}$ moieties, respectively, and two water molecules per unit cell are also found on the 3-fold axes in the large cavity.

Biological Function of Single Chain Glycoprotein Hormone Mutants

  • Min, Kwan-Sik;Chang, Yoo-Min;Chang, Sun-Hwa;Lee, Hyen-Gi;Lee, Yun-Gun;Chang, Won-Kyong;Cheong, Il-Cheong
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.54-54
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    • 2001
  • Human chorionic gonadotropin (hCG) is a member of the glycoprotein hormone family which includes FSH, hCG, TSH. These hormone family is characterized by a heterodimeric structure composed a common $\alpha$-subunit noncovalently linked to a hormone specific $\beta$-subunit. The correct conformation of the heterodimer is also important for efficient secretion, hormone-specific post-translational modifications, receptor binding and signal transduciton. To determine $\alpha$ and $\beta$-subunits can be synthesized as a single polypeptide chain (tethered-hCG) and also display biological activity, the tethered-hCG molecule by fusing the carboxyl terminus of the hCG $\beta$-subunit to the amino terminus of the $\alpha$-subunit was constructed and transfected into chinese hamster ovary (CHO-K1) cells. We also constructed C-terminal deletion mutants (D9l, D89, D88, D87, D86, D84, D83) of single chain hCG to determine the biological function (secretion, LH-activity, receptor binding, cAMP production) of these mutants. Between six and eight stably transfected pools of cells expressing wild type and mutant hCGs were selected for neomycin resistant. The hCGs secreted by the stably transfected cells into serum-free media were collected and quantified by radioimmunoassay, as described in protocol (DPC(hCG IRMA). LH activity was in terms of testosterone production and aromatase activity in primary cultured rat Leydig cells. The tethered-wthCG was efficiently secreted and showed similar LH-like activity to the dimeric hCG. The D83hCG mutant was not detected in this assay. It is suggest that hCG C-terminal part is very important for hCG secretion. Now, we checking the LH-like activity of these mutant hCGs. These data indicate that the constructs of tethered molecule will be useful in the study of mutants that affect subunit association and/or secretion.

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Molecular Conductance Switching Processes through Single Ruthenium Complex Molecules in Self-Assembled Monolayers

  • Seo, So-Hyeon;Lee, Jeong-Hyeon;Bang, Gyeong-Suk;Lee, Hyo-Yeong
    • Proceedings of the Korean Vacuum Society Conference
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    • 2011.02a
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    • pp.27-27
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    • 2011
  • For the design of real applicable molecular devices, current-voltage properties through molecular nanostructures such as metal-molecule-metal junctions (molecular junctions) have been studied extensively. In thiolate monolayers on the gold electrode, the chemical bonding of sulfur to gold and the van der Waals interactions between the alkyl chains of neighboring molecules are important factors in the formation of well-defined monolayers and in the control of the electron transport rate. Charge transport through the molecular junctions depends significantly on the energy levels of molecules relative to the Fermi levels of the contacts and the electronic structure of the molecule. It is important to understand the interfacial electron transport in accordance with the increased film thickness of alkyl chains that are known as an insulating layer, but are required for molecular device fabrication. Thiol-tethered RuII terpyridine complexes were synthesized for a voltage-driven molecular switch and used to understand the switch-on mechanism of the molecular switches of single metal complexes in the solid-state molecular junction in a vacuum. Electrochemical voltammetry and current-voltage (I-V) characteristics are measured to elucidate electron transport processes in the bistable conducting states of single molecular junctions of a molecular switch, Ru(II) terpyridine complexes. (1) On the basis of the Ru-centered electrochemical reaction data, the electron transport rate increases in the mixed self-assembled monolayer (SAM) of Ru(II) terpyridine complexes, indicating strong electronic coupling between the redox center and the substrate, along the molecules. (2) In a low-conducting state before switch-on, I-V characteristics are fitted to a direct tunneling model, and the estimated tunneling decay constant across the Ru(II) terpyridine complex is found to be smaller than that of alkanethiol. (3) The threshold voltages for the switch-on from low- to high-conducting states are identical, corresponding to the electron affinity of the molecules. (4) A high-conducting state after switch-on remains in the reverse voltage sweep, and a linear relationship of the current to the voltage is obtained. These results reveal electron transport paths via the redox centers of the Ru(II) terpyridine complexes, a molecular switch.

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Crystal Structure Theory and Applications of 14-Ethoxy-4,6,-dimethyl-8.12- dioxa-4.6-diazatetracyclo [8.8.0.02,7.013,18]octadeca-13,15,17-triene-3,5-dione

  • Ganapathy, Jagadeesan;Sivakumar, G.;Manickam, Bakthadoss;Sanmargam, Aravindhan
    • Journal of Integrative Natural Science
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    • v.8 no.1
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    • pp.19-29
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    • 2015
  • In view of the growing medicinal importance of chromene and its derivatives, the single crystal X-ray diffraction study was carried out for the potential active 4,6-dimethyl-9-phenyl-8,12-dioxa-4,6-diazatetracyclo [8.8.0.02,7.013,18]octadeca-2(7),13,15,17-tetraene-3,5,11-trione-2-ethoxyphenyl (2E)-but-2-enoate ($C_{18}H_{20}N_2O_5$). In the title compound are two molecules exist in the asymmetric unit. It crystallizes in the monoclinic space group $P2_1/c$ with unit cell dimension a=14.608(3) ${\AA}$, b=12.845(3) and c= 17.781(4) [alpha & gamma=$90^{\circ}$ beta=$91.233(5)^{\circ}$]. Both pyran and pyran ring of the chromene moiety adopts sofa conformation in the molecule A & B. The crystal structure is stabilized by intramolecular C-H...O hydrogen bond interaction.

Conformation of single polymer molecule in a slot coating flow

  • Lee, Jeong-Yong;Ryu, Bo-Kyung;Lee, Joo-Sung;Jung, Hyun-Wook;Hyun, Jae-Chun
    • Korea-Australia Rheology Journal
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    • v.20 no.2
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    • pp.89-94
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    • 2008
  • To satisfy good mechanical and optical properties of polymer-coated film products, it will be indispensable to elucidate the molecular orientation of polymer chains within coating liquids in coating flows. Using hybridized numerical method between computational fluid dynamics (CFD) and Brownian dynamics (BD) simulations can provide the useful information for the better quality control of coated films. Flexible polymer chains, e.g., ${\lambda}$-DNA molecules here, change their conformation according to the flow strength and the flow type. The molecular conformation within the coated film on the web or substrate is quite different, because the polymer chains experience the complicated flow strength and flow types in flow field. Especially in the slot coating flow, these chains are more extended by the extension-like flow field generated in the free surface curvature just beyond the downstream die region. Also, the polymer chain extension beneath the free surface can be affected by the die geometry, e.g., the coating gap, changing flow field.

Investigation of the Binding Force between Protein A and Immunoglobulin G Using Dielectrophoretic(DEP) Tweezers Inside a Microfluidic Chip (미세유체 칩 내에서 유전영동 집게(Dielectrophoretic Tweezers) 를 이용한 단백질A와 면역 글로불린 G의 결합에 관한 연구)

  • Kwak, Tae Joon;Lee, Jae Woo;Yoon, Dae Sung;Lee, Sang Woo
    • Journal of Biomedical Engineering Research
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    • v.34 no.3
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    • pp.123-128
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    • 2013
  • The 'Dielectrophoretic Tweezers(DEP Tweezers)' can be used as a facile, economical toolkit for quantitative measurement of chemical and biological binding forces related to many biological interactions within a microfluidic device. Our experimental setup can probe the interaction between a single receptor molecule and its specific ligand. Immunoglobulin G(IgG) functionalized on polystyrene microspheres has been used to detect individual surface linked Staphylococcus protein A(SpA) molecules and to characterize the strength of the noncovalent IgG-SpA bond. It was measured and compared with the existing measurements. Measured single binding force of between Goat, Rabbit IgG and SpA were $17{\pm}7pN$, $74{\pm}16pN$. This work can be used to investigate several different ligand-receptor interactions and antigen-antibody interactions.

Ultrasonic Relaxation for Complexation Reaction Between β-Cyclodextrin and Butanoic Acid in Aqueous Solution (사이크로덱스트린과 부탄산의 복합체 형성반응에 의한 초음파 완화)

  • Bae, Jong-Rim
    • The Journal of the Acoustical Society of Korea
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    • v.31 no.2
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    • pp.100-106
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    • 2012
  • The aim of this work was to reveal the effect of butanoic acid in its dissociated form and undissociated form as a guest molecule on the kinetic parameters in an inclusion reaction with ${\beta}$-cyclodextrin (${\beta}$-CD). Ultrasonic absorption measurements in the frequency range from 0.2 to 45 MHz were carried out for ${\beta}$-CD solutions with butanoic acid at $25^{\circ}C$ by ultrasonic relaxation method. The rate and the equilibrium constants were obtained from the guest concentration dependence of the relaxation frequency, and the standard volume change of the complexation reaction were obtained from the maximum absorption per wavelength. A single relaxational absorption was observed, and the cause of the relaxation was attributed to a perturbation of the chemical equilibrium associated with a complexation reaction between ${\beta}$-CD and butanoic acid. These results were compared with those in solutions containing both ${\beta}$-CD and different guest molecules. It was found that the hydrophobicity of guest molecules played an important role in the formation of the inclusion complex and also that the charge on the carboxylic group had a considerable effect on the kinetic characteristics of the complexation reaction.

State-selective Dissociation of Water Molecules on MgO Films Using LT-STM

  • Shin, Hyung-Joon;Jung, J.;Motobayashi, K.;Kim, Y.;Kawai, M.
    • Proceedings of the Korean Vacuum Society Conference
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    • 2011.08a
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    • pp.112-112
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    • 2011
  • The interaction of water molecules with solid surfaces has been a subject of considerable interests, due to its importance in the fields from atmospheric and environmental phenomena to biology, catalysis and electrochemistry [1,2]. Among various kinds of surfaces, a lot of theoretical and experimental studies have been performed regarding water on MgO(100), however, to date, there has been no direct observation of water molecules on MgO by scanning tunneling microscope (STM) as compared with those on metal surface. Here, we will present the direct observation and manipulation of single water molecules on ultrathin MgO(100) films using low-temperature scanning tunneling microscope (LT-STM) [3]. Our results rationalize the previous theoretical predictions of isolated water molecules on MgO including the optimum adsorption sites and non-dissociative adsorption of water. Moreover, we were able to dissociate a water molecule by exciting the vibrational mode of water, which is unattainable on metal surfaces. The enhanced residual time of tunneling electrons in molecules on the insulating film is responsible for this unique pathway toward dissociation of water.

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